The kidney is a major site of insulin metabolism removing the hormone from the circulation by means of glomerular filtration and by extraction from the peritubular circulation. Insulin removed binds to specific receptors in the luminal (L) and contraluminal (CL) tubular membranes, the latter is followed by activation of Na-K-ATPase. Filtered insulin is internalized by means of endocytosis and then degraded completely. Whether insulin exposed to the CL membrane undergoes internalization is unknown but CL degradation does occur with formation of partial and complete degradation products. Our specific aims are: I. To determine the ultrastructural pathway and fate of insulin within the kidney: In particular, we aim (a) to resolve the conflict regarding the pathway of filtered insulin after it is absorbed in the proximal tubule, (b) to assess whether insulin extracted from the peritubular circulation is internalized or whether it localizes solely to the cell membrane, and (c) to evaluate the role of lysosomes in renal insulin degradation. II. To evaluate the interaction between insulin and renal tubular plasma membrane receptors. In particular, we aim (a) to determine whether insulin receptors in renal L and CL membranes are regulated in the same manner and whether regulation of these receptors differs from that of receptors in other tissues, (b) to determine whether reversible hyperinsulinuria associated with diabetic ketoacidosis is due to altered renal L membrane insulin binding, (c) to evaluate the role of insulin receptor binding and activation of Na-K-ATPase in the pathogenesis of Na retention following insulin therapy of uncontrolled diabetes. Studies will be conducted with intact rats, isolated perfused rat kidneys and isolated renal plasma membranes. Rats with chemical or spontaneous diabetes will be used. Methodology includes electron microscopic autoradiography, subcellular fractionation, gel filtration, radioimmunoassay, and receptor binding. These studies are particularly significant because of the major role played by the kidney in insulin metabolism. By the use of sophisticated procedures, new information will be generated that should achieve our long term objectives of enhancing the understanding of insulin metabolism, not only in the kidney but also in other organs. In addition, new insight into changes in renal function associated with diabetes will be provided.